In Situ Estimation of Hydrogen Isotope Fractionation Associated with Sucrose and Cellulose Synthesis from Leaves to Roots

Plant cellulose hydrogen (H) stable isotope compositions (δ2H) integrate hydrological and biochemical information, and therefore measurements from archives such as tree rings can be valuable for understanding past climate and plant metabolic responses to environmental change. Although the hydrological component that is integrated into cellulose δ2H values is relatively well understood, the biochemical reactions that can alter δ2H values of metabolites used for cellulose biosynthesis remain cryptic. Attempts at establishing models to simplify the interpretation of cellulose δ2H values have been made, like the widely used cellulose δ2H model by Roden et al. (2000) using the terms quantified by Yakir & DeNiro (1990). However, independent quantification of the parameters in this model, and assessment of their variability with respect to plant C metabolism, has been limited. The cellulose δ2H model uses the δ2H compositions of leaf water and source water, autotrophic and heterotrophic 2H-fractionation (εA and εH, respectively), and the proportion of carbon (C) bound H that exchanges with xylem water during cellulose biosynthesis (ƒ) to explain variation in cellulose δ2H values. By growing plants along a gradient of source water δ2H values under autotrophic and heterotrophic conditions, the original, εA, εH, and ƒ were determined for the aquatic plant Lemna gibba L.. One drawback of this approach is that it assumes these terms are the same when plants are grown in the light vs the dark. We recently reassessed the model for terrestrial plants by measuring δ2H values of leaf sucrose and found species variation in εA (Holloway-Phillips et al., 2022), but were unable to resolve variation associated with ƒ and εH. In the present experiment we assessed a new experimental approach to quantify all model parameters for autotrophically grown plants using regression analysis. This required growing plants with variation in the isotopic offset between xylem water and leaf water (∆LW) and measuring sucrose and cellulose δ2H values from leaves and roots. In a previous study we determined that mutation-induced inhibition of starch synthesis in leaves resulted in higher cellulose δ2H values compared with the wildtype, which was hypothesized to occur preceding sucrose synthesis in the leaves (Baan et al., 2023). Using this new approach, we tested whether this effect was indeed mostly established in source cells during de novo sucrose synthesis (εA), or was a result of 2H-fractionating processes in sink cells prior to cellulose synthesis (εH and ƒ). Preliminary analyses show an increase in leaf sucrose δ2H values in the mutant relative to the wild type, implying that εA is also dependent on plant C metabolism within a given species.